The objective of this research is to develop a model system for the investigation of metabolic diseases in which the enzymatic lesion(s) results in ketonuria. The epitome of these disorders is Maple Syrup Urine Disease, whose pathology stems from defective branched-chain keto acid metabolism. It is proposed to investigate selected areas of metabolism to elucidate the molecular bases of deviate physiology inherent in these diseases. The specific areas include: i. correlation between keto acid imbalances and alterations in cellular lipids ii. relationship between lipid alterations and membrane-related phenomena such as transport and antibiotic sensitivity iii. effects of branched-chain keto acid imbalances upon the metabolism of other keto acids and energy yielding pathways. Pseudomonads will be grown in a defined media in which branched-chain amino acids are the sole carbon source. Gas liquid chromatography will quantitate and qualitate resultant lipid perturbations. Uptake of amino acid and sugar isotopes will determine if chemical differences effect the biological function of cellular lipids. Likewise, antibiotic sensitivity will be monitored to detect changes in resistance. Lipogenesis and branched-chain amino acid catabolic pathways are connected via branched-chain keto acid dehydrogenase which converts keto acids to acyl-Coenzyme A derivatives. This enzyme will be purified and characterized. Mutants blocked in keto acid catabolism and regulatory mutants not subject to branched-chain amino acid inhibitions will be used.